1. Field of the Invention
The present invention relates generally to the fields of developmental biology and molecular biology. More particularly, it concerns an anti-hypertrophic helicase expressed specifically in heart tissue.
2. Description of Related Art
It has been reported by the American Heart Association (1997, Statistical Supplement), that almost 60 million people in the United States suffer from one or more cardiovascular diseases. Cardiovascular diseases are responsible for almost a million deaths annually in the United States representing over 40% of all deaths. Coronary heart disease, characterized by atherosclerotic narrowing of the coronary arteries, resulted in death for almost half a million people in 1997 and is the single leading cause of death in America today. This year it is estimated more than one million Americans will have a new or recurrent coronary attack, and more than 40 percent of the people experiencing these attacks will die of them. Myocardial infarction (MI), commonly referred to as heart attack, is a leading cause of mortality with 30% being fatal in the first months following the attack. Myocardial infarctions result from narrowed or blocked coronary arteries in the heart which starves the heart of needed nutrients and oxygen.
Another form of heart disease, congestive heart failure, represents the most frequent non-elective cause of hospitalization in the U.S. Each year, close to half a million patients are diagnosed with CHF, which is defined as abnormal heart function resulting in inadequate cardiac output for metabolic needs (Braunwald, 1988). Symptoms of CHF include breathlessness, fatigue, weakness, leg swelling, and exercise intolerance. On physical examination, patients with heart failure tend to have elevations in heart and respiratory rates, rales (an indication of fluid in the lungs), edema, jugular venous distension, and, in general, enlarged hearts, indicative of cardiac hypertrophy. Although medical therapy can initially attenuate the symptoms of heart failure (e.g., edema, breathlessness and fluid in the lungs), and in some cases prolong life, the prognosis in this disease, even with medical treatment, is grim (see, e.g., Baughman, 1995). Once symptoms of heart failure are moderately severe, the prognosis is worse than most cancers in that 50% of such patients may die within 2 years (Braunwald, 1988).
Cardiac hypertrophy is an adaptive response of the heart to virtually all forms of cardiac disease, including those arising from hypertension, mechanical load, myocardial infarction, cardiac arrythmias, endocrine disorders and genetic mutations in cardiac contractile protein genes. While the hypertrophic response is initially a compensatory mechanism that augments cardiac output, sustained hypertrophy can lead to dilated cardiomyopathy, heart failure, and sudden death. In the United States, approximately half a million individuals are diagnosed with heart failure each year, with a mortality rate approaching 50%. Because cardiac hypertrophy can be viewed as an aberration in heart growth and development, a relevant inquiry may be made into the molecular basis of cardiac tissue specification and differentiation.
The heart is the first organ to form during mammalian embryogenesis (Olson and Srivastava, 1996; Fishman and Olson, 1997). Formation of the heart involves commitment of cells from the anterior lateral mesoderm to a cardiogenic fate in response to inductive cues from adjacent endoderm. During mouse development, cardiac precursor cells are localized to a region known as the cardiac crescent, which spans the anterior ventral midline of the embryo. These cells migrate ventrolaterally to form a linear heart tube at E8.0. The linear heart tube is patterned along its anterior-posterior axis into segments that give rise to the atria, left ventricle, right ventricle, and outflow tract. Rightward looping of the heart tube is essential for orientation of the right and left ventricular chambers and alignment of the heart with the inflow and outflow tracts. Later events of chamber maturation, septation, endocardial development, and valvulogenesis give rise to the mature multi-chambered heart.
Several mouse and zebrafish mutants exhibit specific defects in cardiac looping, ventricular morphogenesis and chamber maturation (Fishman and Olson, 1997). The phenotypes of these mutants, which often result in ablation of specific segments of the heart, have led to the notion that distinct transcriptional networks control formation of different cardiac compartments. Many of the genes shown to be required for these morphogenetic events encode transcription factors, but the target genes that mediate the actions of these factors are largely unknown.
The basic helix-loop-helix (bHLH) transcription factors, dHAND and eHAND, are expressed specifically in the developing right and left ventricular chambers, respectively. dHAND is required for formation of the left ventricle of the heart (Srivastava et al., 1995, 1997; Firulli et al., 1998; Srivastava, 1999). Similarly, the cardiac homeodomain protein Nkx2.5 is required for looping morphogenesis (Lyons, 1995), and is a regulator of eHAND expression (Biben and Harvey, 1997). The zinc finger transcription factors GATA-4 in mice and GATA-5 in zebrafish have also been shown to be required for ventral morphogenesis and formation of the linear heart tube (Kuo et al., 1997; Molkentin et al., 1997; Reiter et al., 1999).
Recently, the inventors showed that the MADS-box transcription factor MEF2C, which is expressed throughout the linear, looping, and multichambered heart, is required for looping morphogenesis and right ventricular development (Lin et al., 1997). There are four MEF2 genes in vertebrates, MEF2A, -B, -C, and -D, which are expressed in overlapping patterns in developing muscle and neural cell lineages, and at lower levels in other cell types (Black and Olson, 1998). MEF2 factors bind an A/T-rich sequence in the control regions of numerous skeletal, cardiac, and smooth muscle-specific genes. Functional redundancy among the vertebrate MEF2 genes has precluded a complete analysis of MEF2 function in the mouse. However, in Drosophila, there is only one MEF2 gene, which, like the vertebrate MEF2 genes, is expressed in developing muscle cell lineages (Lilly et al., 1994; Nguyen et al., 1994). In Drosophila embryos lacking MEF2, skeletal, cardiac, and visceral myoblasts are properly specified and positioned, but they cannot differentiate, and there are severe abnormalities in morphogenesis of the visceral musculature (Lilly et al., 1995; Ranganayakulu et al., 1995; Bour, 1905). This severe muscle phenotype suggests that MEF2 acts in myoblasts to activate downstream muscle-specific genes involved in differentiation and morphogenesis.
In addition to regulating muscle-specific genes, MEF2 has been implicated in activation of growth factor-inducible and stress-responsive genes (Naya and Olson, 1999). The c-jun promoter, for example, contains a MEF2 site that confers serum and EGF-inducibility (Han et al., 1992, 1995). Signal-dependent activation of MEF2-targeted genes has been shown to involve MAP kinase (Zhao et al., 1999), CaM kinase (Passier et al., 2000), and calcineurin (Chin et al., 1998; Mao et al., 1999). The Notch signaling pathway has been shown to inhibit MEF2 activity in vertebrates and Drosophila (Wilson-Rawls et al., 1999). However, relatively little is know about the targets of MEF2 activation.